title: Contribution of the KCa3.1 channel-calmodulin interactions to the regulation of the KCa3.1 gating process.
abstract: The Ca(2+)-activated potassium channel of intermediate conductance, KCa3.1, is now emerging as a therapeutic target for a large variety of health disorders. The Ca(2+) sensitivity of KCa3.1 is conferred by the Ca(2+)-binding protein calmodulin (CaM), with the CaM C-lobe constitutively bound to an intracellular domain of the channel C terminus. It was proposed on the basis of the crystal structure obtained for the C-terminal region of the rat KCa2.2 channel (rSK2) with CaM that the binding of Ca(2+) to the CaM N-lobe results in CaM interlocking the C-terminal regions of two adjacent KCa3.1 subunits, leading to the formation of a dimeric structure. A study was thus undertaken to identify residues of the CaM N-lobe-KCa3.1 complex that either contribute to the channel activation process or control the channel open probability at saturating Ca(2+) (Pomax). A structural homology model of the KCa3.1-CaM complex was first generated using as template the crystal structure of the C-terminal region of the rat KCa2.2 channel with CaM. This model was confirmed by cross-bridging residues R362 of KCa3.1 and K75 of CaM. Patch-clamp experiments were next performed, demonstrating that the solvation energy of the residue at position 367 in KCa3.1 is a key determinant to the channel Pomax and deactivation time toff. Mutations of residues M368 and Q364 predicted to form anchoring points for CaM binding to KCa3.1 had little impact on either toff or Pomax. Finally, our results show that channel activation depends on electrostatic interactions involving the charged residues R362 and E363, added to a nonpolar energy contribution coming from M368. We conclude that electrostatic interactions involving residues R362 and E363 and hydrophobic effects at M368 play a prominent role in KCa3.1 activation, whereas hydrophobic interactions at S367 are determinant to the stability of the CaM-KCa3.1 complex throughout gating.
authors.affiliation: Department of Physiology, Université de Montréal, Montréal, Québec H3C 3J7, Canada.
authors.forename: Patricia
authors.initials: P
authors.lastname: Morales
authors.forename: Line
authors.initials: L
authors.lastname: Garneau
authors.forename: Hélène
authors.initials: H
authors.lastname: Klein
authors.forename: Marie-France
authors.initials: MF
authors.lastname: Lavoie
authors.forename: Lucie
authors.initials: L
authors.lastname: Parent
authors.forename: Rémy
authors.initials: R
authors.lastname: Sauvé
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002147:Calmodulin / Q000737:chemistry* / Q000378:metabolism; D051660:Intermediate-Conductance Calcium-Activated Potassium Channels / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D015640:Ion Channel Gating; D056004:Molecular Dynamics Simulation; D008969:Molecular Sequence Data; D009154:Mutation; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D021122:Protein Subunits / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D051381:Rats; D055672:Static Electricity
chemical_list: D002147:Calmodulin; D051660:Intermediate-Conductance Calcium-Activated Potassium Channels; C491575:Kcnn4 protein, rat; D021122:Protein Subunits; D002118:Calcium